The enteric nervous system (ENS), the largest and most phenotypically diverse region of the PNS, has unique characteristics, which are reminiscent of the brain, and are not seen in other ganglia. We have found that the neural crest-derived precursors of enteric neurons probably begin to differentiate as neurons before they reach the bowes; however, the microenvironment provided by the non-neuronal cells of the wall of the gut may be important in determining what type of neuron these precursors will form and also whether other precursors will express the phenotype of enteric glia or Schwann cells. The goal of the current proposal is to test the hypothesis that the development of the nuerons and gial of the ENS, as well as its unique pattern of organization, results from a determinative interaction between neural crest emigres and the enteric mesenchyme. In vitro experiments, in which the conditions of this interaction can be controlled, will be done to test this hypothesis and determine the nature of responsible factors. Quail neural crest, which can be recognized and traced, will be combined in co-culture with chick enteric mesenchyme. Intermediate filament content, neurotransmitter-related properties, and ultrastructure will be used as markers of the enteric pattern of ganglionic development. Developmental regulation, regional specificity and the role of soluble factors, extracellular matrix and/or cell to cell contact in the neural crest-enteric mesenchyme interaction will be explored. The development of the progeny of single clones of neural crest cells in the enteric mesenchyme will be followed to evaluate the extent to which single migrating neural crest cells are multipotential. Regions of the neuraxis that do or do not normally provide cells for the ENS will be compared. Finally, the regulation of specific transient changes in the enteric mesenchyme that may be associated with the development of particular types of enteric neuron will be analyzed. These changes include the specific uptake of serotonin by mesenchymal cells and the expression of aspects of a catecholaminergic phenotype by proliferating cells in the enteric mesenchyme that may be neuroblasts. Analysis of transient monoaminergic expression will use cell culture, in situ hybridization with a radiolabeled cDNA probe for tyrosin hydroxylase mRNA and the type-specific cellular localization of monamine oxidase. Selection of the serotonergic neuronal phenotype and the possibility that ingrowing sympathetic nerves regulate gene expression by enteric neuroblasts will be investigated.